The Drosophila undergoes a complete metamorphosis in its progression from an embryo to an adult. The embryo is characterized by its polarity, which readily distinguishes the anterior and posterior, dorsal and ventral parts of the animal, and by serially repeating segments (or metameres), each with characteristic structural patterns. The adult body is also segmented, and groups of these segments are organized into a head, thorax, and abdomen.
Three major classes of pattern-regulating genes specify the basic features of the Drosophila embryo's body and function in successive states of development. The maternal genes are expressed in the unfertilized egg and provide most of the proteins needed in very early development. The maternal genes direct the spatial organization of the developing embryo at early stages, establishing its polarity. The segmentation genes are expressed after fertilization and direct the formation of the proper number of body segments. Finally, the homeotic genes are expressed later and affect the unique characteristics of the individual body segments.
In general, the homeotic genes control the fundamental architectural plan of the embryo. The two main clusters of homeotic genes found in the Drosophila genome are the Antennapedia complex (ANT-C) and the Bithorax complex (BX-C). The ANT-C directs the proper development of the head and anterior thoracic segments during fruitfly development and comprises five homeotic genes in the following sequential order: labial (lab), proboscipedia (pb), Deformed (Dfd), Sex combs reduced (Scr), and Antennapedia (Antp). The BX-C directs the proper development of the posterior thoracic segments and abdominal segments and comprises three homeotic genes in the following sequential order: Ultrabithorax (Ubx), abdominal A (abdA), and Abdominal B (AbdB). All of these homeotic genes code for specific RNA polymerase II transcription factors which are required for DNA transcription. Thus, the homeotic genes regulate segment development and body plan formation by regulating DNA transcription.
The genetic order of the homeotic genes within the two complexes correlates with the sequence of metameres on which they act. Thus, the most proximate member of the ANT-C complex, the labial (lab) gene, is the most anteriorly expressed and is critical for the development of the embryonic and adult head. In contrast, the AbdB gene of the BX-C complex is the most posteriorly expressed homeotic gene and affects the formation of structures at the posterior end of the embryo or the adult fly. Thus, the functional specificity of the different homeotic genes is achieved partly by their precise spatial and temporal expression patterns.
It has been reported that two of the Drosophila homeotic genes, Antp and Ubx, are translated by a unique cap-independent mechanism in contrast to the cap-dependent mechanism utilized by most other eukaryotic genes. Eukaryotic mRNAs have a distinctive structural feature at its 5′ end, called a 5′ cap, which is a residue of 7-methylguanosine linked to the 5′ terminal residue of the mRNA through an unusual 5′, 5′-triphosphate linkage. Cap-dependent translation is initiated by the binding of the cap-binding protein complex eIF-4F to the 5′ cap, which in turn facilitates the binding of the 43S ternary ribosomal subunit near or at the 5′ cap region. The ribosome complex is purported to scan the mRNA from the 5′ cap until it encounters the first AUG initiation codon, where translation of the mRNA is initiated. (see Kozak, M, (1989) Cell 44:283–292; Kozak, M (1989) J. Cell. Biol. 108:229–241).
The cap-independent translation mechanism was proposed to explain the efficient translation of some mRNAs despite the presence of a highly ordered RNA structure in the 5′ untranslated region (5′UTR) of mRNAs which was predicted to interfere with ribosome scanning of the mRNA. The picornavirus mRNA was the first mRNA identified that displayed a cap-independent translation mechanism. The picornavirus mRNA is characterized by a unique structure, including the absence of a 5′ cap, the presence of an extraordinarily long and structured 5′ UTR, and the presence of multiple upstream AUG initiation codons. This long and structured 5′UTR was found to serve as an internal ribosome entry site (IRES) or a ribosome landing pad, where the 43S ternary ribosomal subunit would bind and initiate translation independently of the 5′ cap structure.
The 5′UTR containing an IRES is generally characterized by three complex features: a long 5′UTR, a stable secondary structure, and potential upstream AUG initiation codons. The stable secondary structure is considered to be the major determinant of IRES function. A low proportion of vertebrate mRNAs have long, highly structured 5′UTRs that contain multiple AUG initiation codons. Among these, the Drosophila Antp gene has been found to harbor a 1,735 nt-long 5′UTR and 15 upstream AUG codons, and the Ubx gene has a 968 nt-long 5′UTR and two upstream AUG codons. To date, a limited, but a growing subset of IRESs have been identified in cellular mRNAs in various species including human (Macajak, D. G. and P. Sarnow, (1991) Nature 353:653–656; Sarnow, P, (1989) PNAS 86:5795–5799; Vagner, S. et al., (1995) Mol. Cell. Biol. 15:35–44), and yeast (Zhou, W. et al., (2001) PNAS 98:1531–1536; Paz, I. et al., (1999) J. Biol. Chem. 274:21741–21745). IRESs have also been identified in viral mRNAs, such as in poliovirus (Pelletier, J. and N. Sonenberg. (1988) Nature 334:320–325), encephalomyocarditis virus (EMCV) (Jang, S. K., and E. Wimmer, (1990) Genes Dev. 4:1560–1572), and human rhinovirus (HRV) (Borman, A. et al., (1993) J. Gen. Virol. 74:1775–1788). The Antp and Ubx homeotic genes of Drosophila are also translated via an IRES in their long 5′UTRs (Ye X. et al., (1997) Mol. Cell. Biol. 17:1714–1721; Ho, S.-K. et al., (1992) Genes Dev. 6:1643–1653).